Accumulation of intracellular HCO3- by Na+-HCO3- cotransport in interlobular ducts from guinea-pig pancreas
Accumulation of intracellular HCO3- by Na+-HCO3- cotransport in interlobular ducts from guinea-pig pancreas
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DOI：
10.1113/jphysiol.1996.sp021582
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发表时间：
1996-08-15
影响因子：
5.5
通讯作者：
Case, RM
中科院分区：
文献类型：
--
作者：
Ishiguro, H;Steward, MC;Case, RM
1. 酶法消化后，从豚鼠胰腺上显微解剖小叶间管的短段。过夜培养后，用微荧光法测量细胞内pH（pH(i)）和Na+浓度（[Na+](i)），这些细胞在装载pH敏感的荧光探针2'7'-双（2-羧基乙基）-5(6)-羧基荧光素（BCECF）或钠结合苯并呋喃异眼酸钠（SBPI） 2的导管细胞中。通过使用NH4Cl脉冲技术对细胞进行酸负荷，研究了负责维持pH(i)高于平衡的转运蛋白。在没有HCO3-/CO2的情况下，pH(i)的恢复依赖于Na+，被0.2 mM的阿米洛林和10 μ的n -甲基- n -异丁胺洛林所消除，因此归因于Na+- h +交换。在HCO3-/CO2存在下，阿米洛利仅部分抑制酸负荷的回收。对酰胺不敏感的组分被0.5 mM H2DIDS消除，不受细胞内Cl-消耗的影响，因此归因于Na+- hco3 -共转运。4. 尽管HCO3-外排显著增加，但10 nM分泌素刺激并未引起pH(i)的显著变化。然而，在分泌素存在的情况下，添加0.5 mM H2DIDS导致pH(i)的下降速度是0.2 mM amiloride的3倍。在分泌素刺激的导管中，当向培养液中加入HCO3-/CO2时，Na+的摄取增加，这种增加被0.5 mM h2did强烈抑制。我们得出结论，在分泌素刺激下，Na+-HCO3-共转运贡献了豚鼠胰管细胞约75%的HCO3-吸收。有人提出，腔膜上的HCO3-外排和基底膜上的电致Na+-HCO3-共运输之间的电偶联解释了分泌素导致pH变化很小的原因(i)。
1. Short segments of interlobular duct were microdissected from guinea-pig pancreas following enzymatic digestion. After overnight culture, intracellular pH (pH(i)) and Na+ concentration ([Na+](i)) were measured by microfluorometry in duct cells loaded with either the pH-sensitive fluoroprobe 2'7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF) or the sodium-binding benzofuran isophthalate (SBPI).2. The transporters responsible for maintaining pH(i) above equilibrium were investigated by using the NH4Cl pulse technique to acid load the cells. In the absence of HCO3-/CO2, the recovery of pH(i) was Na+ dependent, abolished by 0.2 mM amiloride and by 10 mu N-methyl-N-isobutylamiloride and was therefore attributed to Na+-H+ exchange.3. In the presence of HCO3-/CO2, amiloride only partially inhibited the recovery from acid loading. The amiloride-insensitive component was abolished by 0.5 mM H2DIDS and unaffected by depletion of intracellular Cl- and was therefore attributed to Na+-HCO3- cotransport. 4. Stimulation with 10 nM secretin did not cause a significant change in pH(i) despite a significant increase in HCO3- efflux. However, in the presence of secretin, addition of 0.5 mM H2DIDS caused a decline in pH(i) that was three times more rapid than that obtained with 0.2 mM amiloride.5. In secretin-stimulated ducts, Na+ uptake increased when HCO3-/CO2 was added to the bath and this increase was strongly inhibited by 0.5 mM H2DIDS.6. We conclude that Na+-HCO3- cotransport contributes approximately 75% of the HCO3- taken up by guinea-pig pancreatic duct cells during stimulation with secretin. It is proposed that electrical coupling between HCO3- efflux at the luminal membrane and electrogenic Na+-HCO3- cotransport at the basolateral membrane explains why secretin causes little change in pH(i).